Abstract
Alpha-synuclein (Snca) plays a major role in Parkinson disease (PD). Circulating anti-Snca antibodies has been described in PD patients and healthy controls, but they have been poorly characterized. This study was designed to assess the prevalence of anti-Snca reactivity in human subjects carrying the LRRK2 mutation, idiopathic PD (iPD) patients, and healthy controls and to map the epitopes of the anti-Snca antibodies. Antibodies to Snca were detected by ELISA and immunoblotting using purified recombinant Snca in plasma from individuals carrying LRRK2 mutations (104), iPD patients (59), and healthy controls (83). Epitopes of antibodies were mapped using recombinant protein constructs comprising different regions of Snca. Clear positive anti-Snca reactivity showed no correlation with age, sex, years of evolution, or the disability scores for PD patients and anti-Snca reactivity was not prevalent in human patients with other neurological or autoimmune diseases. Thirteen of the positive individuals were carriers of LRRK2 mutations either non-manifesting (8 out 49 screened) or manifesting (5 positive out 55), three positive (out of 59) were iPD patients, and five positive (out of 83) were healthy controls. Epitope mapping showed that antibodies against the N-terminal (a.a. 1–60) or C-terminal (a.a. 109–140) regions of Snca predominate in LRRK2 mutation carriers and iPD patients, being N122 a critical amino acid for recognition by the anti-C-terminal directed antibodies. Anti-Snca circulating antibodies seem to cluster within families carrying the LRRK2 mutation indicating possible genetic or common environmental factors in the generation of anti-Snca antibodies. These results suggest that case-controls’ studies are insufficient and further studies in family cohorts of patients and healthy controls should be undertaken, to progress in the understanding of the possible relationship of anti-Snca antibodies and PD pathology.
Highlights
Parkinson disease (PD) and other synucleinopathies are characterized pathologically by proteinaceous inclusions commonly described as Lewy pathology in postmortem brain tissue samples (Goedert, 2001)
Snca has been found in cerebrospinal fluid (CSF) and plasma and its concentration in these fluids has been suggested as a possible biomarker for PD (Parnetti et al, 2013), even though there are substantial discrepancies between the results reported in different studies (Lee et al, 2006; Li et al, 2007; Duran et al, 2010; Shi et al, 2010; Mollenhauer et al, 2011; Park et al, 2011; Foulds et al, 2012; Wennstrom et al, 2013)
Www.frontiersin.org positive samples by immunoblot, we found that some of those clearly positive by ELISA did not recognize the Snca protein itself, but minor, likely E. coli, protein contaminants that were not obvious by Coomassie blue staining. These inconsistent results made us to introduce a further step of purification of Snca by RP-HPLC, rending a preparation of Snca close to 98% homogenous
Summary
Parkinson disease (PD) and other synucleinopathies (dementia with Lewy bodies and multiple system atrophy) are characterized pathologically by proteinaceous inclusions commonly described as Lewy pathology in postmortem brain tissue samples (Goedert, 2001). Snca has been found in cerebrospinal fluid (CSF) and plasma and its concentration in these fluids has been suggested as a possible biomarker for PD (Parnetti et al, 2013), even though there are substantial discrepancies between the results reported in different studies (Lee et al, 2006; Li et al, 2007; Duran et al, 2010; Shi et al, 2010; Mollenhauer et al, 2011; Park et al, 2011; Foulds et al, 2012; Wennstrom et al, 2013). The extracellular behavior of Snca is part of the rationale for an immunological intervention as Frontiers in Aging Neuroscience www.frontiersin.org
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